Gene regulation in response to heat stress

响应热应激的基因调控

• 批准号: 1518345
• 负责人: David Gross
• 金额: $ 51万
• 依托单位: LSU Health Sciences Center -Shreveport
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-01 至 2019-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1518345&HistoricalAwards=false
• 关键词: Gene; regulation; response; heat; stress

This project will investigate the mechanisms that allow organisms to survive after exposure to high temperatures. Using baker's yeast (Saccharomyces cerevisiae), experiments will focus on understanding how cells coordinate genome-wide changes in gene expression following heat stress. Because the heat stress response is evolutionarily conserved, research using yeast as the model is expected to provide insights into the fundamental mechanisms employed in more complex organisms. Additional broader impacts of this research include the training of two graduate students and summer research opportunities for three high school biology teachers from a school district with a large, underrepresented minority population. The dynamic regulation of stress-responsive genes is critical for viability of all eukaryotes. In S. cerevisiae, the gene-specific and evolutionarily conserved activator, Heat Shock Factor 1 (Hsf1), plays a central role in stimulating both basal and induced transcription of HSP genes. These genes encode molecular chaperones that are critical for maintaining protein homeostasis and in combating proteotoxic stress. Whether Hsf1 is also responsible for the global down-regulation of non-HSP gene expression following exposure to thermal stress is unknown. Aim 1 employs chromatin immunoprecipitation combined with deep sequencing (ChIP-seq) to investigate genome-wide occupancy of Hsf1 under both non-stressful and stressful conditions, and determine the activator's role in directing the genome-wide occupancy of Mediator, a central coactivator and signal integrator of Pol II transcription. In addition, comparative dynamic transcriptome analysis will be used to measure global transcription rates and correlate these with genome-wide Hsf1, Mediator and Pol II occupancy. Aim 2 employs chromosome conformation capture techniques to characterize the in vivo conformation and nuclear organization of Hsf1-regulated genes under both non-inducing and heat shock-inducing conditions. A key goal is to solidify evidence that HSP genes loop, 'crumple' (accordion-style) and coalesce into transcriptionally active foci following exposure of cells to heat shock, and determine the kinetics with which they form and dissipate looped DNA structures. Additionally, through use of looping, crumpling and coalescence-deficient mutants, the biological significance of these phenomena will be explored. Results are expected to advance understanding of how gene regulatory changes enable cell survival following heat stress.

这个项目将研究使生物体在高温下存活的机制。使用面包酵母（Saccharomyces cerevisiae），实验将重点了解细胞如何协调热应激后基因表达的全基因组变化。由于热应激反应在进化上是保守的，使用酵母作为模型的研究有望为更复杂的生物体所采用的基本机制提供见解。这项研究的其他更广泛的影响包括培训两名研究生，并为来自一个拥有大量未被充分代表的少数民族人口的学区的三名高中生物教师提供暑期研究机会。应激反应基因的动态调控对真核生物的生存至关重要。在酿酒酵母中，基因特异性和进化保守的激活因子热休克因子1 （Hsf1）在刺激HSP基因的基础和诱导转录中起着核心作用。这些基因编码的分子伴侣对于维持蛋白质稳态和对抗蛋白质毒性应激至关重要。Hsf1是否也对暴露于热应激后非hsp基因表达的全局下调负责尚不清楚。目的1采用染色质免疫沉淀结合深度测序（ChIP-seq）研究Hsf1在非应激和应激条件下的全基因组占用，并确定激活子在指导Mediator （Pol II转录的中心辅助激活子和信号整合子）的全基因组占用中的作用。此外，比较动态转录组分析将用于测量全球转录率，并将其与全基因组Hsf1、Mediator和Pol II占用率相关联。目的2采用染色体构象捕获技术来表征hsf1调控基因在非诱导和热休克诱导条件下的体内构象和核组织。一个关键的目标是巩固证据，证明热休克基因在细胞暴露于热休克后形成环状，“皱褶”（手风琴式）并合并成转录活性灶，并确定它们形成和消散环状DNA结构的动力学。此外，通过使用环，皱缩和聚结缺陷突变体，将探索这些现象的生物学意义。研究结果有望促进对热应激后基因调控变化如何使细胞存活的理解。